Alzheimer's disease is progressive dementia occurring at the presenile stage (between the ages of 45 and 65). It causes morbid changes such as degeneration of neurons and atrophy of cerebral cortex due to a decrease in the number of neurons. Pathologically, a number of senile plaques and neurofibrillary degeneration are observed in the brain. So-called senile dementia caused by spontaneous aging in the senium at the age of 65 or older is not substantially different from Alzheimer's disease from the pathological viewpoint and is regarded as senile dementia of the Alzheimer type. This disease has been perceived as a social problem because the number of patients suffering from Alzheimer's disease increases as the senile population increases. Although there are various hypotheses about the causes of this disease, it remains to be elucidated and an early breakthrough in clarification of the disease is desired.
The main component of senile plaques that is one of the pathological changes caused by Alzheimer's disease is known to be amyloid β protein (Annu. Rev. Neurosci., 12, 463-490 (1989)). Neurofibrillary degeneration that is another pathological change shows accumulation of the paired helical filament (may hereinafter be referred to as PHF) in neurons and phosphorylated tau protein is identified as one of its constituents (J. Biochem. 99, 1807-1810 (1986); Proc. Natl. Acad. Sci. USA, 83, 4913-4917 (1986)).
Although tau protein is composed of a group of protein isoforms that usually form several bands at a molecular weight of 48 to 65 kD (as a result of SDS-polyacrylamide gel electrophoresis) and promotes formation of microtubules, tau protein incorporated in the PHF of the Alzheimer diseased brain has been proven to be abnormally phosphorylated as compared with that in the normal brain using a polyclonal antibody to PHF (anti-p-tau; J. Biochem., 99, 1807-1810 (1986)) and a monoclonal antibody to tau protein (tau-1 antibody; Proc. Natl. Acad. Sci. USA, 83, 4913-4917 (1986)). Further, the phosphorylation sites of phosphorylated tau protein incorporated in the PHF have been identified (JP 6-239893 A), and functions of tau protein involved in Alzheimer's disease is now being clarified.
For tau proteins, a kit for measuring the concentration of a tau protein in cerebrospinal fluid (trade name in Japan “Finoscolor hTAU” and trade name in the U.S. and Europe “INNOTEST hTAU Ag”, manufactured by Innogenetics) has been already commercially available and clinically utilized. A method of detecting Alzheimer's disease based on the phosphorylation site of a phosphorylated tau protein in PHF has been developed (Neurosci. Lett., 270, 91-94 (1999); Ann. Neurol., 50, 150-156 (2001)). Both of those methods are designed to use cerebrospinal fluid as a sample. The collection of the samples is problematic for patients due to its highly invasive nature. Thus, in light of in the need to reduce invasiveness for patients and convenient analysis, there have been strong demands for specific and sensitive methods of detecting Alzheimer's disease in which a CNS tau protein can be analyzed even by using a variety of samples not limited to cerebrospinal fluid and including blood.
However, it has been revealed that several different isoforms exist for the tau protein. A central nervous tissue, such as the brain, and a peripheral tissue, such as the muscle, have different isoforms of the tau protein which predominantly exist in each tissue (J. Neurochem., 67, 1235-1244 (1996)). For example, the isoform of a tau protein that predominantly exists in a peripheral tissue (hereinafter, which is also referred to as a “peripheral tau protein”) has a large molecular weight as compared to the isoform of a tau protein that predominantly exists in a central nervous tissue (hereinafter, which is also referred to as a “CNS tau protein”). It has been suggested that, in a blood sample or the like, this peripheral tau protein having a large molecular weight is contained in a large amount.
The inventors of the present invention have previously proposed a method of detecting Alzheimer's disease using blood as a sample (JP 2002-040023 A). However, in this method, the different isoforms of a tau protein are not distinguished, and thereby all are detected. Therefore, the method does not allow one to specifically analyze a change in the CNS tau protein that occurs in the brain of a patient with Alzheimer's disease. For example, when a blood sample or the like is analyzed using such a method, the detection of a CNS tau protein is interfered with by the strong reaction of a peripheral tau protein having a large molecular weight that is also contained in a large amount in the blood sample. As a result, sufficient sensitivity was not obtained.
In short, it has been difficult to analyze a tau protein derived from a central nervous tissue whose content is low in a blood sample, or the like in which extremely high concentrations of gross proteins are present resulting in a large interference due to impurities (Dement. Geriat. Cong. Disord., 10, 442-445 (1999); Neurosci. Lett., 275, 159-162 (1999)). Thus, the establishment of specific and sensitive methods of detecting Alzheimer's disease has not been established.
In addition, mild cognitive impairment (hereinafter, which may be abbreviated to “MCI”) showing a subjective symptom of memory loss or the like has recently received attention as prodrome of Alzheimer's disease. Of patients diagnosed as having MCI, 10-15% in a year and 50% in several years have been said to proceed to Alzheimer's disease, and there is a growing acknowledgement that patients with early Alzheimer's disease are included in MCI patients. However, existing evaluation for MCI complies with a criterion such as the criteria of Petersen, R. C. et al., (Arch. Neurol., 56, 303-308 (1999)) that focuses on history taking or intelligent function examinations. Thus, objective and clear diagnostics have not been established. Furthermore, those conventional methods are unable to detect patients with early Alzheimer's disease among a group of patients diagnosed as having MCI. Consequently, there is a strong demand to establish methods of distinctly detecting Alzheimer's disease for patients including such patients diagnosed as having MCI.